Antimicrobial Activity of Ceftazidime-Avibactam and Comparators against Pathogens Harboring OXA-48 and AmpC Alone or in Combination with Other β-Lactamases Collected from Phase 3 Clinical Trials and an International Surveillance Program

ABSTRACT In vitro activities of ceftazidime-avibactam (CAZ-AVI) and key comparators against AmpC-overproducing Enterobacterales and Pseudomonas aeruginosa isolates from four Phase 3 clinical trials and against OXA-48–producing Enterobacterales with multiple resistance mechanisms from the Antimicrobial Testing Leadership and Surveillance (ATLAS) program were evaluated. Susceptibility to CAZ-AVI and comparators was determined by reference broth microdilution methods. Clinical response at test of cure (TOC) was assessed in patients from Phase 3 trials with baseline OXA-48–producing Enterobacterales or AmpC-overproducing Enterobacterales and P. aeruginosa treated with CAZ-AVI or comparators. Against 77 AmpC-overproducing Enterobacterales isolates from Phase 3 trials, meropenem-vaborbactam (98.7% susceptible [S]), CAZ-AVI (96.1% S), and meropenem (96.1% S) had similar in vitro activity and were more active than ceftolozane-tazobactam (24.7% S). Clinical cure rates in patients with baseline AmpC-overproducing Enterobacterales were 80.7% (n = 21/26) and 85.0% (n = 17/20) for CAZ-AVI and comparators. Against 53 AmpC-overproducing P. aeruginosa isolates from Phase 3 trials, CAZ-AVI (73.6% S) was more active in vitro than ceftolozane-tazobactam (58.5% S) and meropenem (37.7% S). Clinical cure rates in patients with baseline AmpC-overproducing P. aeruginosa were 85.7% (n = 12/14) and 75.0% (n = 9/12) for CAZ-AVI and comparators, respectively. Of 113 OXA-48–producing isolates from the ATLAS program, 99.1% were susceptible to CAZ-AVI. Four patients with baseline OXA-48–producing Klebsiella pneumoniae isolates treated with CAZ-AVI in Phase 3 trials were clinical cures at TOC and had favorable microbiological response. CAZ-AVI was among the most active agents against AmpC-overproducing P. aeruginosa and Enterobacterales and had greater in vitro activity against OXA-48–producing Enterobacterales than meropenem-vaborbactam, meropenem, ceftolozane-tazobactam, and other comparators.

carbapenems (3,4). OXA-48-producing isolates, predominantly Klebsiella pneumoniae, are increasingly prevalent in many parts of the world and are dominant in certain regions, such as North Africa, the Middle East, and Europe (1,4,5). A major contributor of the dissemination is the acquisition of plasmid-mediated bla OXA-48 genes. OXA-48 is inhibited by avibactam, but not inhibited by traditional b-lactamase inhibitors and other non-b-lactam-based inhibitors. Ceftazidime is not significantly hydrolyzed by OXA-48 but potentially can lose activity in the presence of ESBLs (1,5).
AmpC b-lactamases, on the other hand, are more complex owing to inducible expression in variable levels of the chromosomal encoded bla ampC gene (cAmpC), as well as the constitutively expressed plasmid-encoded bla ampC genes (pAmpC) (6). A stable derepression of bla ampC expression often results in the upregulation of expression of the AmpC b-lactamases, which in turn decreases the susceptibility of cephalosporins in pathogens that harbor the inducible system (6). Strains producing cAmpC in an inducible manner usually appear susceptible in in vitro assays to third-generation cephalosporins, which are weak inducers but can constitutively produce the enzymes that cause resistance to these drugs, resulting in treatment failures (2,6). Pathogens coproducing AmpC and ESBLs may exhibit multidrug-resistant phenotypes, limiting treatment options owing to coexpression of multiple resistance determinants. AmpCs are usually resistant to b-lactam-based b-lactamase inhibitors but are well inhibited by non-b-lactam-based inhibitors, such as avibactam (6).
This study evaluated in vitro activities of CAZ-AVI and key comparators against AmpC-overproducing Enterobacterales and P. aeruginosa isolates collected from Phase 3 clinical trials, RECLAIM (10), RECAPTURE (11), REPROVE (12), and REPRISE (13) ( Table 1) and OXA-48-producing Enterobacterales from surveillance studies. RECLAIM, RECAPTURE, and REPROVE were double-blind, noninferiority studies that evaluated the efficacy, safety, and tolerability of CAZ-AVI in the treatment of hospitalized adults with complicated intra-abdominal infections (cIAI), complicated urinary tract infections (cUTI), or hospital-acquired pneumonia/ventilator-associated pneumonia (HAP/VAP) caused by Gram-negative pathogens. REPRISE was an open-label clinical study that evaluated the efficacy of CAZ-AVI compared with best available therapy (primarily carbapenems) in patients with either cIAI or cUTI caused by ceftazidime-resistant Gram-negative pathogens. OXA-48-producing Enterobacterales with multiple resistance mechanisms were also collected from a CAZ-AVI global surveillance program (ATLAS: Antimicrobial Testing Leadership and Surveillance), and susceptibility of these isolates to CAZ-AVI and comparator antibiotics was tested as part of this study. Finally, clinical response at test of cure (TOC) was also assessed for patients from the Phase 3 studies that had AmpC-overproducing or OXA-48-producing Enterobacterales and P. aeruginosa at baseline.  Table 2).

DISCUSSION
The novel b-lactam-b-lactamase inhibitor combinations (CAZ-AVI, ceftolozane-tazobactam, and meropenem-vaborbactam) are a significant advance in the therapeutic armamentarium against multidrug-resistant Gram-negative pathogens. The CAZ-AVI combination has shown potent activity against carbapenemase-producing Enterobacterales and P. aeruginosa owing to inhibitory activity of avibactam toward carbapenemases, including OXA-48 and KPC (2,6). Moreover, avibactam also inhibits ESBLs and Class C cephalosporinases, offering a viable treatment option for infections caused by pathogens carrying OXA-48 or AmpC alone or in combination with ESBLs. Although the TABLE 5 In vitro susceptibility of CAZ-AVI and comparator agents tested against Enterobacterales producing AmpC alone or in combination with other resistance mechanisms a b-lactam-b-lactamase inhibitor combinations are highly effective against large collections of clinical isolates, each has a unique susceptibility profile (14). While tazobactam does not inhibit bla OXA-48 , activity of ceftolozane-tazobactam may be expected against isolates with OXA-48-like enzymes, which are poorly active against ceftolozane and other oxyimino-cephalosporins. However, a study showed that ceftolozane-tazobactam displayed limited in vitro activity against Enterobacterales isolates harboring OXA-48. In a study of 353 OXA-48-producing isolates, despite the presence of tazobactam, which should inhibit ESBLs, ceftolozane-tazobactam MICs closely tracked those of unprotected ceftazidime: 81.9% of ceftazidime-susceptible/intermediate isolates (ceftazidime MIC #4 mg/L, EUCAST criteria) were susceptible to ceftolozane-tazobactam, with only 8.1% of ceftazidime-resistant isolates (ceftazidime MIC . 4 mg/L, EUCAST criteria) reported as susceptible to ceftolozane-tazobactam (15). The explanations include the possibility of OXA-48 overwhelming tazobactam, rendering it unable to protect ceftolozane against the AmpC or ESBL enzymes also present in these isolates, or the carriage of additional porin mutations. Vaborbactam, a cyclic boronic acid BLI, exhibited no activity against isolates harboring bla OXA-48 when combined with meropenem (5). Previously published in vitro susceptibility data of MVB showed that the combination had no or limited activity against bla OXA-48 carrying Enterobacterales (5,16). In this study, MVB exhibited moderate to high activity against these isolates, significantly higher than that of MEM alone with blaOXA-48 alone or in combination with other b-lactamases. However, the MIC 50 and MIC 90 of MVB were identical to those of MEM (2/32 mg/L), which is consistent with observations that vaborbactam does not inhibit bla OXA-48 . The difference in susceptibility can be attributed to a higher breakpoint set for MVB (susceptible #4 mg/L) than that for MEM alone (susceptible #1 mg/L). An efficacy study of human-simulated exposure of MVB and MEM in the neutropenic murine thigh infection model revealed that the activity of both drugs against bla OXA-48 carrying Enterobacterales was poor despite more than a third of isolates falling within the susceptible range per EUCAST and CLSI MIC interpretation criteria (8). Therefore, caution needs to be taken when interpreting in vitro susceptibility data for bla OXA-48 carrying Enterobacterales to ensure successful clinical outcomes. In addition, vaborbactam is a potent inhibitor of Class A b-lactamases (17); however, for Enterobacterales carrying bla OXA-48 in combination with CTX-M, SHV, TEM, etc., this study showed that the susceptibility of MVB was dramatically lower than that of isolates carrying bla OXA-48 alone. Enterobacterales showed a relatively high susceptibility (80%) to AMK. The activity of AMK against isolates carrying bla OXA-48 in various combinations with ESBLs and other b-lactamases decreased by approximately 2-10%. As the presence of carbapenems or b-lactamases is not expected to impact aminoglycoside susceptibility, aminoglycoside-modifying genes, 16S rRNA methylases, and other potential mechanisms remain to be investigated. This in vitro susceptibility study demonstrated that CAZ-AVI was the most active agent in vitro against OXA-48producing Enterobacterales carrying multiple b-lactamases compared with other antibiotics, including b-lactams, b-lactam-b-lactamase inhibitor combinations, and aminoglycosides.
The clinical trial data review demonstrated successful clinical outcomes for patients treated with CAZ-AVI who had infections caused by OXA-48-producing organisms, although the number of patients was small. In another observational study evaluating 57 patients receiving CAZ-AVI treatment for infections caused by OXA-48-producing Enterobacterales, CAZ-AVI showed promising results, even in monotherapy, for the treatment of patients with severe infections due to OXA-48-producing Enterobacterales and limited therapeutic options (18).
AmpC is known to be inhibited by non-b-lactam-based inhibitors, such as avibactam, although there may be some variability in susceptibility to inhibitors (2,6). In this study, CAZ-AVI demonstrated strong in vitro activity against Enterobacterales and P. aeruginosa isolates overproducing AmpC b-lactamase along with ESBLs, whereas for ceftazidime, like other cephalosporins, the activity is compromised, resulting in limited in vitro activities against the same isolates. Moreover, ceftolozane-tazobactam is much less potent against the Enterobacterales strains overproducing AmpC b-lactamase and ESBLs owing to the weak inhibitory activity of tazobactam against inducible and constitutively expressed AmpC enzymes (18). Nevertheless, against P. aeruginosa-overproducing AmpC b-lactamase and ESBLs, the in vitro susceptibility to ceftolozane-tazobactam was improved.
In CAZ-AVI regulatory clinical trials evaluating the efficacy of CAZ-AVI and carbapenem comparators, the clinical success rates in patients with baseline AmpC-overproducing P. aeruginosa were 86% (n = 12/14) in the CAZ-AVI group versus 75% (n = 9/12) in the carbapenem control groups. Clinical success rates in patients with baseline AmpCoverproducing Enterobacterales were 81% (n = 21/26) in the CAZ-AVI group versus 85% (n = 17/20) in carbapenem groups. Similar results were observed in a systematic review and meta-analysis of randomized controlled trials comparing the clinical efficacy of CAZ-AVI and carbapenems for the treatment of ESBL/AmpC-producing Enterobacterales; the clinical response for AmpC producers in CAZ-AVI and carbapenem arms was 80% (n = 32/40) and 88% (n = 37/42), respectively (2). These results showed evidence of the clinical efficacy of ceftazidime-avibactam as a potential alternative to carbapenems in patients with AmpC-overproducing or OXA-48-producing Enterobacteriaceae and P. aeruginosa. In a retrospective multicenter study evaluating clinical success in patients hospitalized in 13 Italian hospitals who received $72 h of CAZ-AVI, a 90% clinical cure rate was observed in all 41 assessed patients at the end of CAZ-AVI treatment despite the study population having a higher prevalence of infections caused by MDR, XDR, and PDR, including OXA-48, AmpC, ESBL, and KPC pathogens (19).
In this descriptive analysis, the number of isolates at baseline overproducing AmpC b-lactamase or OXA-48 carbapenemase was small, limiting interpretation of the clinical outcome evaluation. Additional limitations are that the data were pooled from four clinical trials that had differences in design, limiting direct comparison. As a result, the analyses carried out were exploratory, and the results should be interpreted with caution.
In conclusion, our study showed that CAZ-AVI was among the most active agents against AmpC-overproducing P. aeruginosa and Enterobacterales and had greater in vitro activity against OXA-48-producing Enterobacterales than comparators.
Resistant subsets. b-Lactamase gene screening in AmpC-overproducing clinical trial isolates was conducted using either microarrays (Check-Points, Wageningen, Netherlands) and PCR or a combination of both as described previously (20)(21)(22)(23). The presence and expression levels of AmpC were determined using quantitative PCR, and the threshold of upregulation was 5-fold above a reference value as described previously (22). The screening was conducted at central reference laboratories (JMI, North Liberty, IA, USA).
OXA-48-producing Enterobacterales isolates were screened for genes encoding carbapenemases (KPC, OXA-48-like, NDM, IMP, VIM) as well as the presence of coharbored b-lactamase genes encoding TEM, SHV, CTX-M-1 group, CTX-M-2 group, CTX-M-8 group, CTX-M-9 group, CTX-M-25 group, ACC, ACT, CMY, and DHA using a combination of microarray and multiplex PCR assays, followed by amplification and sequencing of the full-length genes. The screening was conducted at the central reference laboratory (IHMA, Schaumburg, IL, USA).